Engineering Bacteriophytochrome-coupled Photoactivated Adenylyl Cyclases for Enhanced Optogenetic cAMP Modulation

• Published in: Journal of molecular biology Vol. 436; no. 5; p. 168257
• Main Authors: Xu, Qianzhao, Vogt, Arend, Frechen, Fabian, Yi, Chengwei, Küçükerden, Melike, Ngum, Neville, Sitjà-Roqueta, Laia, Greiner, Andreas, Parri, Rhein, Masana, Mercè, Wenger, Nikolaus, Wachten, Dagmar, Möglich, Andreas
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier Ltd 01.03.2024
• Subjects: cyclic nucleotide; optogenetics; phytochrome; second messenger; sensory photoreceptor; phytochrome; second messenger; cyclic nucleotide; optogenetics; sensory photoreceptor
• ISSN: 0022-2836; 1089-8638; 1089-8638
• DOI: 10.1016/j.jmb.2023.168257

[Display omitted] •Bacteriophytochromes (BphP) regulate effector function by red and far-red light.•Recombining BphPs and cyclases generates photoactivated adenylyl cyclases (PAC)•The variant DmPAC exhibits 40-fold enhanced cyclase activity under red light.•The so-called PHY tongue governs PAC activity and light response.•DmPAC affords optogenetic control of cyclic-nucleotide levels in mammalian cells. Sensory photoreceptors abound in nature and enable organisms to adapt behavior, development, and physiology to environmental light. In optogenetics, photoreceptors allow spatiotemporally precise, reversible, and non-invasive control by light of cellular processes. Notwithstanding the development of numerous optogenetic circuits, an unmet demand exists for efficient systems sensitive to red light, given its superior penetration of biological tissue. Bacteriophytochrome photoreceptors sense the ratio of red and far-red light to regulate the activity of enzymatic effector modules. The recombination of bacteriophytochrome photosensor modules with cyclase effectors underlies photoactivated adenylyl cyclases (PAC) that catalyze the synthesis of the ubiquitous second messenger 3′, 5′-cyclic adenosine monophosphate (cAMP). Via homologous exchanges of the photosensor unit, we devised novel PACs, with the variant DmPAC exhibiting 40-fold activation of cyclase activity under red light, thus surpassing previous red-light-responsive PACs. Modifications of the PHY tongue modulated the responses to red and far-red light. Exchanges of the cyclase effector offer an avenue to further enhancing PACs but require optimization of the linker to the photosensor. DmPAC and a derivative for 3′, 5′-cyclic guanosine monophosphate allow the manipulation of cyclic-nucleotide-dependent processes in mammalian cells by red light. Taken together, we advance the optogenetic control of second-messenger signaling and provide insight into the signaling and design of bacteriophytochrome receptors.